Abstract:
:When yeast are faced with irreparable DNA damage, they will first arrest in G2/M, via the DNA damage checkpoint pathway, but will subsequently adapt to that arrest and resume division. Here, we summarize assays that we have used to examine checkpoint adaptation. Specifically, we discuss the merits of inducing DNA damage with ionizing radiation (IR) and IR-mimetic drugs, HO, and the cdc13-1 mutation. We also discuss readouts that we have used to visualize adaptation.
journal_name
Methods Enzymoljournal_title
Methods in enzymologyauthors
Toczyski DPdoi
10.1016/S0076-6879(05)09009-9subject
Has Abstractpub_date
2006-01-01 00:00:00pages
150-65eissn
0076-6879issn
1557-7988pii
S0076-6879(05)09009-9journal_volume
409pub_type
杂志文章abstract::In this chapter, we motivate the need for photoactivatable NO donor molecules and give a brief survey of the existing chemical tools in the field. We then provide detailed protocols for the synthesis and validation of a near-infrared light-activated NO donor molecule, photoNOD-1, developed in our research group. With ...
journal_title:Methods in enzymology
pub_type: 杂志文章
doi:10.1016/bs.mie.2020.05.003
更新日期:2020-01-01 00:00:00
abstract::Slp4-a/granuphilin-a is a member of the synaptotagmin-like protein (Slp) family and consists of an N-terminal Slp homology domain (SHD) and C-terminal tandem C2 domains. Slp4-a is specifically localized on secretory granules in some endocrine and exocrine cells through its SHD, and it attenuates Ca(2+)-dependent dense...
journal_title:Methods in enzymology
pub_type: 杂志文章
doi:10.1016/S0076-6879(05)03039-9
更新日期:2005-01-01 00:00:00
abstract::Posttranslational modifications (PTMs) of proteins perform crucial roles in regulating the biology of the cell. PTMs are enzymatic, covalent chemical modifications of proteins that typically occur after the translation of mRNAs. These modifications are relevant because they can potentially change a protein's physical ...
journal_title:Methods in enzymology
pub_type: 杂志文章,评审
doi:10.1016/S0076-6879(09)63040-8
更新日期:2009-01-01 00:00:00
abstract::The methods of computational chemistry and molecular modeling are becoming more and more accessible to biochemists with the advent of fast, inexpensive graphics workstations and well-tested computer programs. The state of the art in small molecules allows chemists to use these programs as "black boxes" and be confiden...
journal_title:Methods in enzymology
pub_type: 杂志文章
doi:10.1016/0076-6879(94)43042-x
更新日期:1994-01-01 00:00:00
abstract::Granzyme B (GrB) is the primary molecular mediator of apoptosis by cytotoxic T lymphocytes (CTLs) and natural killer (NK) cells. It is a unique mammalian aspartic acid-cleaving serine protease. On T cell receptor activation, GrB is released from the CTL cytoplasmic granules by exocytosis, enters the target cells and, ...
journal_title:Methods in enzymology
pub_type: 杂志文章
doi:10.1016/s0076-6879(00)22013-2
更新日期:2000-01-01 00:00:00
abstract::Cholestatic syndromes are inborn or acquired disorders of bile formation. In recent years, several inherited cholestatic syndromes were characterized at the molecular level: progressive familial intrahepatic cholestasis (PFIC) and benign recurrent intrahepatic cholestasis (BRIC). Both PFIC and BRIC were divided phenot...
journal_title:Methods in enzymology
pub_type: 杂志文章,评审
doi:10.1016/S0076-6879(05)00031-5
更新日期:2005-01-01 00:00:00
abstract::The mapping and characterization of the vasculature using proteomics offers the opportunity to better understand the steps and molecular mechanisms involved in vascular development, and angiogenesis in particular. Proteomics has many key advantages over genomics, especially in directly determining protein expression. ...
journal_title:Methods in enzymology
pub_type: 杂志文章
doi:10.1016/S0076-6879(08)03008-5
更新日期:2008-01-01 00:00:00
abstract::Here we describe an optimized recombineering protocol for the creation of conditional alleles. Recombineering is a DNA engineering methodology based on homologous recombination in Escherichia coli. It presents a number of advantages over conventional engineering methods, including the ability to place target sites for...
journal_title:Methods in enzymology
pub_type: 杂志文章
doi:10.1016/S0076-6879(10)77008-7
更新日期:2010-01-01 00:00:00
abstract::Epitope tagging has provided a useful experimental strategy with widespread applicability. The ample variety of epitope tag systems that have been put to use to date provide a collection of attributes relevant to virtually any experimental system. As a consequence, epitope tagging will continue to be a valuable tool f...
journal_title:Methods in enzymology
pub_type: 杂志文章,评审
doi:10.1016/s0076-6879(00)27263-7
更新日期:2000-01-01 00:00:00
abstract::At least two discrete deoxyribonuclease activities can be detected during apoptotic death, one that generates 30- to 500-kilobase pair (kbp) domain-sized fragments and another that mediates internucleosomal DNA degradation. The latter nuclease has been identified as the caspase-activated deoxyribonuclease (CAD)/CPAN, ...
journal_title:Methods in enzymology
pub_type: 杂志文章
doi:10.1016/s0076-6879(00)22003-x
更新日期:2000-01-01 00:00:00
abstract::Cytoskeletal motors utilize the energy stored in ATP to generate linear motion along rigid filaments. Because their enzymatic cycles are tightly coupled to the production of force and forward movement, the optical-trapping technique is uniquely suited for studying their mechanochemical cycle. Here, we discuss the prac...
journal_title:Methods in enzymology
pub_type: 杂志文章
doi:10.1016/bs.mie.2016.09.037
更新日期:2017-01-01 00:00:00
abstract::Biotinylated molecules are extensively employed in bioanalytics and biotechnology. The currently available assays for quantification of biotin groups suffer from low sensitivity, low accuracy, or provide highly variable responses for different biotin derivatives. We developed a competitive binding assay in which avidi...
journal_title:Methods in enzymology
pub_type: 杂志文章
doi:10.1016/bs.mie.2019.10.028
更新日期:2020-01-01 00:00:00
abstract::Proteins change their charge state through protonation and redox reactions as well as through binding charged ligands. The free energy of these reactions is dominated by solvation and electrostatic energies and modulated by protein conformational relaxation in response to the ionization state changes. Although computa...
journal_title:Methods in enzymology
pub_type: 杂志文章,评审
doi:10.1016/bs.mie.2016.05.052
更新日期:2016-01-01 00:00:00
abstract::Patient-derived xenografts are a useful tool in cancer immunology, as they allow researchers to study human cancers in vivo when starting with a relatively small amount of human tumor tissue. These models make it possible to study tumor cell-intrinsic changes that occur in response to external stimuli including cytoki...
journal_title:Methods in enzymology
pub_type: 杂志文章
doi:10.1016/bs.mie.2019.10.027
更新日期:2020-01-01 00:00:00
abstract::Omics approaches have become popular in biology as powerful discovery tools, and currently gain in interest for diagnostic applications. Establishing the accurate genome sequence of any organism is easy, but the outcome of its annotation by means of automatic pipelines remains imprecise. Some protein-encoding genes ma...
journal_title:Methods in enzymology
pub_type: 杂志文章
doi:10.1016/bs.mie.2016.09.019
更新日期:2017-01-01 00:00:00
abstract::The goal of the signal transduction pathways, such as those controlled by Ras, is in large part to ensure highly stringent regulation of the target genes in the nucleus, which are collectively responsible for the signal output, or phenotypes, of the cell. Understanding of the Ras effect ultimately requires the identif...
journal_title:Methods in enzymology
pub_type: 杂志文章
doi:10.1016/s0076-6879(01)32206-1
更新日期:2001-01-01 00:00:00
abstract::Undiluted cytoplasmic extract prepared from unfertilized Xenopus laevis eggs by low-speed centrifugation (CSF extracts) is useful for reconstitution of egg microtubule dynamics and meiosis-II spindle organization, but it suffers limitations for biochemical analysis due to abundant particulates. Here, we describe prepa...
journal_title:Methods in enzymology
pub_type: 杂志文章
doi:10.1016/B978-0-12-397924-7.00023-6
更新日期:2014-01-01 00:00:00
abstract::The Arc (anoxic redox control) two-component system (TCS) is a complex signal transduction system that plays an important role in regulating energy metabolism at the level of transcription in bacteria. This system comprises the ArcB protein, a hybrid membrane-associated sensor kinase, and the ArcA protein, a typical r...
journal_title:Methods in enzymology
pub_type: 杂志文章
doi:10.1016/S0076-6879(10)71012-0
更新日期:2010-01-01 00:00:00
abstract::Proteins are fragile molecules that often require great care during purification to ensure that they remain intact and fully active. Nowadays, many proteins are also purified in small amounts under denaturing conditions by various gel electrophoretic techniques, such that inactive proteins are obtained. But even here,...
journal_title:Methods in enzymology
pub_type: 杂志文章,评审
doi:10.1016/S0076-6879(09)63010-X
更新日期:2009-01-01 00:00:00
abstract::The steady technical advances of nuclear magnetic resonance (NMR) over the past decades enabled a significant increase in the molecular size of protein particles that can be subjected to a structural and functional characterization in solution. The larger molecular weight of such proteins is accompanied with an increa...
journal_title:Methods in enzymology
pub_type: 杂志文章
doi:10.1016/bs.mie.2015.05.028
更新日期:2015-01-01 00:00:00
abstract::Peptides corresponding to the calmodulin-binding domain of MLCK and other target enzymes promise to be useful tools in the study of calmodulin action. Not only will they help to elucidate the molecular interactions of calmodulin with specific target enzymes, but they should also prove to be useful calmodulin antagonis...
journal_title:Methods in enzymology
pub_type: 杂志文章
doi:10.1016/0076-6879(87)39079-2
更新日期:1987-01-01 00:00:00
abstract::There are estimated to be >300,000 plant species, producing >200,000 metabolites. Many of these metabolites are restricted to specific plant lineages and are referred to as "specialized" metabolites. These serve varied functions in plants including defense against biotic and abiotic stresses, plant-plant and plant-mic...
journal_title:Methods in enzymology
pub_type: 杂志文章
doi:10.1016/bs.mie.2016.02.023
更新日期:2016-01-01 00:00:00
abstract::There is a great need for practicing scientists to volunteer their time and expertise in the K-12th grade science classroom. We have found that bioluminescence is a fun and exciting way to teach basic science concepts and is an excellent tool for the volunteering scientist. We have had very positive reactions from bot...
journal_title:Methods in enzymology
pub_type: 杂志文章
doi:10.1016/s0076-6879(00)05517-8
更新日期:2000-01-01 00:00:00
abstract::Glycoproteins are a major class of glycoconjugates displaying a variety of mutual interactions between glycan and protein moieties that ultimately affect molecular organization. Modulation of the pendant glycan structures is important in tuning the functions of glycoproteins. Here we discuss structural aspects and som...
journal_title:Methods in enzymology
pub_type: 杂志文章
doi:10.1016/S0076-6879(10)78018-6
更新日期:2010-01-01 00:00:00
abstract::The circadian clock exists to synchronize inner physiology with the external world, allowing life to anticipate and adapt to the continual changes that occur in an organism's environment. The clock architecture is highly conserved, present in almost all major branches of life. Within eukaryotes, the filamentous fungus...
journal_title:Methods in enzymology
pub_type: 杂志文章,评审
doi:10.1016/bs.mie.2014.10.009
更新日期:2015-01-01 00:00:00
abstract::Gene manipulation serves the purpose of providing a better understanding of the function of specific genes as well as for developing novel variants of the genes of interest. The generation of knockout genes, the alteration, depletion, or enhancement of a particular gene function through the generation of specific gene...
journal_title:Methods in enzymology
pub_type: 杂志文章
doi:10.1016/B978-0-12-420067-8.00008-8
更新日期:2013-01-01 00:00:00
abstract::Microarrays of defined glycans represent a high throughput approach to determining the specificity of lectins, or more generally glycan-binding proteins (GBPs). The utility of a glycan microarray is directly related to the number and variety of the glycans available on the printed surface for interrogation by GBPs. Th...
journal_title:Methods in enzymology
pub_type: 杂志文章,评审
doi:10.1016/S0076-6879(10)80033-3
更新日期:2010-01-01 00:00:00
abstract::Carbohydrate microarrays are becoming a standard tool for glycobiologists to screen large numbers of sugars and elucidate the role of carbohydrates in biological systems. This article describes detailed methods to prepare and use microarrays containing synthetic oligosaccharides as well as a summary of the biological ...
journal_title:Methods in enzymology
pub_type: 杂志文章
doi:10.1016/S0076-6879(06)15017-X
更新日期:2006-01-01 00:00:00
abstract::RNA and DNA carry out diverse functions in biology including catalysis, splicing, gene regulation, and storage of genetic information. Interest has grown in understanding how nucleic acids perform such sophisticated functions given their limited molecular repertoire. RNA can fold into diverse shapes that often perturb...
journal_title:Methods in enzymology
pub_type: 杂志文章
doi:10.1016/B978-0-12-801122-5.00009-X
更新日期:2014-01-01 00:00:00
abstract::Signal transduction by vascular endothelial growth factors (VEGFs) through their cognate VEGF receptor tyrosine kinases follows the consensus scheme for receptor tyrosine kinases. Thus, binding of ligand induces receptor dimerization and activation of the tyrosine kinase through transphosphorylation between receptor m...
journal_title:Methods in enzymology
pub_type: 杂志文章
doi:10.1016/S0076-6879(08)02013-2
更新日期:2008-01-01 00:00:00